A striking feature of Neisseria gonorrhoea, the causative agent of gonorrhea, is the ability of gonococci to vary expression of key surface components. The Opacity (Opa) outer membrane proteins are subject to such variation. Gonococci may express up to 10 antigenically distinct Opa proteins, each of which shows on-off phase variation. Opa proteins may facilitate adherence/invasion of host cells, and there is strong selection for Opa expression in vivo. The long-term goal of this project is to understand the role of Opa proteins in gonococcal pathogenesis and immunity. The Specific Aims are to address these questions; 1) Is expression of Opa proteins required for human infection? A gonococcal mutant that is unable to express any Opa proteins will be constructed and tested for infectivity in the human challenge model of gonorrhea. Additional experiments will determine if different Opa proteins are functionally equivalent in infection. 2) What is the topology of Opa proteins in the outer membrane? Are any conserved Opa domains exposed on the gonococcal surface? Reporter epitopes will be inserted at different locations throughout the length of apr proteins; surface exposure of the reporters will be assessed by antibody binding to intact bacteria. Chemical modification of surface-exposed amino acids will also be used. 3) Are anti-Opa antibodies associated with protection from pelvic inflammatory disease (PID), as recent studies suggest? If so, are such antibodies directed at conserved or variable domains of the proteins? Sera that were previously obtained from African women in two large cohort studies will be analyzed for antibodies binding to well-characterized recombinant Opa proteins. Epitopes recognized by anti-Opa antibodies will be mapped; the distribution of those epitopes on Opa proteins of strains causing PID will be determined. These studies will begin to clarify the role of Opa proteins in gonococcal infection, and will aid in evaluating the possibility that an anti-Opa immune response might be protective against the most serious complications of gonorrhea in women.